Hereditary mutations in the DNA helicases XPB and XPD lead to human diseases with different phenotypes reflecting increased cancers or aging: xeroderma pigmentosum (XP), XP combined with Cockayne syndrome (CS), and trichothiodystrophy (TTD). These diseases reflect the disruption of different cellular pathways: nucleotide-excision repair (NER), transcription-coupled repair (TCR), or transcription. In humans, XPB and XPD helicases are part of the ten subunit TFIIH transcription/repair complex, but disease-causing mutations cluster in XPB and particularly XPD rather than in the other TFIIH proteins, excepting TFB5, so these XP helicases appear key to controlling coordination of transcription and repair. We aim to understand the molecular features underlying the specificity, activity, conformational controls and pathway coordination by the XPB and XPD helicases. Our hypothesis is that well-defined architectures, conformational states, and molecular interfaces of XPB and XPD helicases provide critical controls for transcription, NER, and TCR. We have shown that characterizations of these features and their disruption by disease-causing mutations provide a molecular basis to directly connect the inherited gene mutations to disease phenotypes. Building on our crystal structures of XPB and XPD, we propose to integrate structural and biophysical experiments including small angle x-ray scattering to define conformations and complexes in solution with biochemical and biological experiments to determine structures of disease-relevant mutants, protein-DNA complexes, and define key interactions for their activities. The anticipated outcome of the proposed cross-disciplinary experiments is a molecular picture of the protein-DNA complexes, protein-protein interactions and functional states that orchestrate transcription and repair events mediated by XPB and XPD as components of TFIIH. These results will help provide a detailed molecular understanding of the processes that underlie the cancer and cell death disease phenotypes associated with XP, XP/CS, and TTD patient mutations. PUBLIC HEALTH RELEVANCE: The XP helicases sit at the crossroads of cancer and aging. As members of the TFIIH complex, they participate in both DNA repair that maintains the integrity of the genetic code, and transcription that executes that code. Defects in this machinery, as small as a single amino acid change, have consequences for human health. The XP helicases are named for their role in Xeroderma pigmentosum (XP) a disease characterized by such extreme skin cancer predisposition that children with this disease are called "children of the moon." Although XP is the most common, defects in XPB and XPD, along with a handful of other genes, cause two other diseases that do not have increased cancer risk. Cockayne Syndrome (CS) and trichothiodystrophy (TTD) are premature aging diseases with profound neurological defects. The goal of this research is to understand the molecular basis for the normal XP helicase activities and for their breakdown in human disease phenotypes associated with defects in the XP helicases.